The alpha22 gene of herpes simplex virus 1 encodes two mRNAs. These direct the synthesis of the 420 amino acid infected cell protein No. 22 (ICP22) and the US 1.5 protein initiated at the ICP22 met147. To facilitate discussion, we define the ICP22 domain as unique to that protein and the US1.5 domain as that shared by the two proteins. Both domains are essential for viral replication in experimental animals but not in HEp-2 or Vero (permissive) cells. The US1.5 domain is necessary for optimal replication in primary human cell cultures and rodent (restrictive) cells. The US1.5 domain is phosphorylated by the UL13 protein kinase,- a requirement for viral replication in experimental animals or for optimal replication in restrictive cells. The ICP22 domain is nucleotidylylated and binds cell cycle-dependent protein p78. A host protein, p60, binds unprocessed US1.5 protein and ICP0. In permissive cells, p60 is translocated to small nuclear bodies with ICP0 even in the absence of US1.5 protein whereas in restrictive cells p60 is posttranslationally processed by ICP22/US1.5 and UL13 proteins but is not translocated. The objectives of this proposal are to (i) define the sequences and separate the functional domains of ICP22 and of US1.5 protein. The fundamental hypothesis is that ICP22/US1.5 gene complex expresses several functions which may be separable into different genes. If true, it would facilitate further studies of their functions; (ii) determine the function of p60 in relation to viral replication in permissive and restrictive cells. The fundamental hypothesis is that p60 must be sequestered in the small nuclear bodies by ICP0 (permissive cells) or posttranslationally modified by ICP22/US1.5 and UL13 proteins (restrictive cells) for optimal replication; (iii) map the site(s) of posttranslational modifications of ICP22 and US1.5 protein. Mutants lacking UL13 exhibit a phenotype similar to that of mutants lacking US1.5 sequences. Is UL13 required solely to phosphorylate US1.5 or does it act independently? (iv) map and define the functions of the ICP22 domain. The hypothesis is that the functions of ICP22 domain are different from those of the US1.5 domain.